The effects of duration of infection and of temperature and nourishment in cultures on development of free-living generations of Strongyloides stercoralis were studied quantitatively. Rhabditiform larvae, 228–353 μm long, were collected from infected dogs with or without prednisolone treatment using the Baermann apparatus. Cultures were carried out by the filter paper test-tube method under the following conditions: incubation temperature 15–40°C and faecal dilution 1:0–1:16. Rhabditiform larvae developed predominantly to free-living females at incubation temperatures of 15–30°C and low faecal dilutions, but filariform larvae appeared mainly under extreme conditions such as high temperature. Recovery rates of filariform larvae were inversely related to those of females. It was remarkable that high temperature, but not low faecal dilution, affected development of filariform larvae. Although the appearance of free-living males was constant in various environmental conditions, the present study indicated an increase in free-living males with the duration of infection. Thus, it seems that free-living males are already fixed as male in the egg stage, and potential female rhabditiform larvae differentiate into free-living females or filariform larvae depending on environmental factors. There is no marked difference in the development of rhabditiform larvae into filariform larvae in either the immunosuppressed dog or the intact dog.

The composition and structure of a community of Eimeria was investigated in a population of Eastern cottontail (Sylvilagus floridanus) introduced into Italy. Eight Eimeria species were found, of which all but 1 had North American origins and were presumably introduced into Italy together with imported cottontails. The success of cottontails in spreading microparasites is probably related to their massive release for hunting purposes. Nearly all cottontails were infected with at least 1 Eimeria species, with bimonthly prevalence ranging from 0–6·3% (E. leporis) to 42·9–89·3% (E. environ). Bayesian model averaging and multivariate techniques were used to investigate the relationships between the occurrence of each parasite and the structure of the relative community. Among the host parameters, only sex was found to be associated with the prevalence of E. honessi, while the rest of the parameters were only weakly correlated with prevalence and species richness. This indicates that individual phenotypic host characteristics are probably less important than environmental factors in determining levels of parasite prevalence and diversity. The community of Eimeria species was probably structured by competition, with less species co-occurrence than expected under a null hypothesis. This was made evident by the low co-occurrence of E. environ and E. neoirresidua with E. poudrei, E. honessi, and E. maior.

This study was conducted to investigate the level of cardiac and oxidative stress markers in camels infected with Trypanosoma evansi and to explore the diagnostic and prognostic value of cardiac troponin I (cTnI) and creatine kinase-myocardial band (CK-MB) in response to infection. Seventy four dromedary camels with clinical and laboratory evidence of trypanosomosis and 20 healthy controls were included in this study. Serum cTnI, CK-MB, CK, malondialdehyde (MDA) and super oxide dismutase (SOD) were measured. The values of cTnI, CK-MB, CK and MDA were significantly higher, whereas SOD level was lower in T. evansi infected camel. Successfully treated camels (n = 43) had lower levels of cTnI, CK-MB, CK and MDA, but higher level of SOD compared to camels with treatment failure. Both cTnI and CK-MB showed high degree of accuracy in predicting treatment outcome (success vs failure). The area under the curve for cTnI and CK-MB was 0·98 and 0·93, respectively. However, cTnI showed better sensitivity and specificity than CK-MB (Se = 96·8% vs 83·9% and Sp = 100% vs 88·5%, respectively). These results suggest that cTnI and CK-MB could be used as diagnostic and prognostic biomarkers in camels infected with T. evansi.

Dientamoeba fragilis is a pathogen of the human gastrointestinal tract that is a common cause of diarrhoea. A paucity of knowledge on the in vitro cultivation and cryopreservation of Dientamoeba has meant that few studies have been conducted to investigate its biology. The objective of this study was to define, for the first time, in vitro culture conditions able to support the long-term in vitro growth of Dientamoeba. Also, we aimed to define a suitable method for cryopreserving viable Dientamoeba trophozoites. A modified BD medium, TYGM-9, Loeffler's slope medium, Robinson's medium, Medium 199, Trichosel and a Tritrichomonas fetus medium were compared, using cell counts, for their ability to support the growth of D. fragilis at various temperatures and atmospheric conditions. Loeffler's slope medium supported significantly better growth compared to other media. A temperature of 42°C and a microaerophilic atmosphere were also optimum for Dientamoeba growth. To our knowledge, this is the first study to describe and compare different culture media and conditions for the growth of clinical isolates of D. fragilis. This new technology will aid the development of diagnostics for dientamoebiasis as well as facilitate large-scale sequencing projects that will fast track molecular studies on D. fragilis.

When eggs of Ascaris lumbricoides Lin. or A. suilla Duj. containing mature embryos gain entrance to the alimentary canal of the sewer rat, Mus decumanus, or the mouse, M. musculus, they hatch. It is not possible to say at present in what part of the alimentary canal hatching takes place. (The number of animals suitable for experiment at the disposal of the writer is unfortunately so small that he is unable to devote any of them to the working out of the details of the process.) A certain proportion of the larvae thus liberated escape in the faeces where under suitable circumstances they can live for at least three days. It is, however, probable they ultimately succumb and that this is not a true road of development.

The majority of the larvae gain entrance into the body of the host. The exact point of entrance and the time after hatching at which entrance takes place have not been determined. Some animals show signs of illness on the second day after infection. The time elapsing between infection and the entrance of the larvae into the body is therefore probably not more than two days.

Larvae are found in the lungs and liver of the host not later than four days after infection and possibly as early as two days. Sections of the tissues show that they are situated in the air vesicles of the lung and in the blood capillaries of the liver close to the interlobular branches of the portal vein.

Larvae are not found in the liver after the fifth day from infection. They are found in the bronchi about the seventh day and in the trachea on the eighth day.

No larvae are found in any portion of the lung on the ninth day after infection. Dead larvae have been found in the stomach and rectum on the ninth day after the last infection.

The route by which the larvae reach these sites is of course not definitely proved but from anatomical considerations it is hardly possible that it can be other than one of the two following (the diameter of the larva is three times that of a red blood corpuscle of the mouse. The larva could therefore not pass through the lumen of an ordinary capillary vessel):

(1) Boring through the wall of the stomach or intestine the larva enters a mesenteric venule and is carried to the liver. It is here arrested at the entrance to the hepatic capillary plexus and it is for this reason that so many larvae are found in the capillaries close to the interlobular veins. The liver undergoes extreme and acute fatty degeneration so that the larvae are able to penetrate along the capillaries between the degenerated columns of liver cells to the hepatic venules. Thence they pass in the hepatic vein to the heart and by the pulmonary artery to the lung. They are of course at once arrested by the pulmonary capillary field. Embolism of the smaller branches of the pulmonary artery takes place with haemorrhage around these arterioles. The larvae readily work their way along with the effused blood into the air vesicles and thence into the bronchi and trachea.

(2) The larva after hatching in the stomach or duodenum travels up the bile duct and reaches the bile capillaries of the interlobular zone. It here bores its way through the degenerated liver tissues and reaching a hepatic venule continues its course as in the first case.

During the residence in the body of the rat or mouse the larvae grow from a length of 0·22 mm. to 1·4 mm. The proportion length of oesophagus/total length diminishes from 1/2·5 to 1/6·1. The ventral line which is the greatest of the longitudinal lines in the embryo is reduced to the same dimensions as the dorsal and lateral lines. The ventral gland (the rudiment of the excretory system) is developed from a cell of the ventral line, enlarges very greatly, acquires the massive nucleus characteristic of Ascaris larvae (Stewart, 1906; Baylis, 1916) and finally develops its duct from the cells of the ventral line. The intestine, anal canal and anus become pervious. The rudiment of the female gonads appears.

Malaria and helminth infections have a shared geographical distribution and therefore co-infections are frequent in tropical areas of the world. Human populations of helminth and malaria co-infection have shown contradictory results for the course of malarial infection and disease, possibly depending on the type of helminth studied, the intensity of helminth infection and the age of the study population. Although immunological studies might clarify the underlying mechanisms of protection or increased susceptibility, there are very few studies that have looked at immunological parameters in helminth and malaria co-infection. After discussing the available immunological data on co-infection, we describe a pilot study performed in Ghanaian school children where we compare anti-malarial responses in children living in an urban area, where the prevalence of helminth and Plasmodium falciparum infections was low, with that of children living in a rural area with high prevalence of helminth and Plasmodium falciparum infections.

The purpose of this work is to review the progress towards malaria vaccination that has been made over the last four or so years, and the prospects and difficulties as they now appear. Although some of the older literature will be referred to as necessary background, it is not treated here in any detail. The reader who wishes for a fuller historical perspective should see, for instance, Brown (1969), Cohen & Mitchell (1978), Desowitz & Miller (1980), Mitchell (1984), Miller, David & Hadley (1984), Heidrich (1986) and, specifically for a consideration of sporozoite vaccination, Nussenzweig & Nussenzweig (1984, 1986). Naturally, any summary of vaccination will lean on the immunology of the disease, but malaria immunology is not reviewed here in its own right. The reader requiring the most recent attempt to cover this field should see the work edited by Perlmann & Wigzell (1988); some individual chapters of that volume will be referred to below.

Rohdella siamensis, a new genus and species of aspidogastrean, is described from freshwater fishes in Thailand. A new subfamily, the Rohdellinae, is erected to accommodate this worm on the grounds that it differs from all known aspidogastreans in that the male and female ducts unite to form an hermaphroditic duct, with the result that the intromittent organ is a sinus-organ and not a cirrus. A new classification of the subclass Aspidogastrea is presented. Two orders are recognized: the Aspidogastrida, containing a single family, the Aspidogastridae, which in turn contains 3 subfamilies, the Aspidogastrinae, the Cotylaspidinae and the Rohdellinae subfam.nov.; and the Stychocotylida ord.nov., containing 3 families, the Stichocotylidae, the Multicalycidae fam.nov. and the Rugogastridae. The systematic position of the Stichocotylida, and especially the Rugogastridae, is commented upon.

‘Each organism must be examined as a biochemical entity before any reasonable understanding of helminth metabolism can be attained’ (Saz, 1969).

The pathogenic flagellate Blastocrithidia triatomae disrupts the digestion of Triatoma infestans; the midgut ultrastructure of bugs infected with the flagellate and of uninfected bugs is compared. Third or fourth instar larvae were dissected either unfed or 1 week after feeding.In all uninfected bugs extracellular membrane layers (e.m.l.) covered the apical microvillar border of the epithelial cells. Some midgut regions of bugs infected with B. triatomae appeared normal but often adjacent cells showed pathological effects. In affected cells the e.m.l. and the microvilli and finally the cells themselves were reduced or destroyed. Correlated with these observations of pathogenicity the method of attachment of parasites changed. When the e.m.l. were present only rarely were flagella found, but on extracellular membrane-free cells B. triatomae attached by flagellar enlargement to the microvillar border or, if this was reduced, to the apical host cell membrane. No hemidesmosome-like plaques were found at the attachment site. Although some flagella were inserted into the apical region of the cells no intracellular flagellates were observed.

An enzyme presenting kallikrein-like activity (designated sK1) was purified from the supernatant of Schistosoma mansoni adult worm homogenate. The enzyme cleaves bradykinin from purified rat plasma kininogen. Activity was optimal at pH 9·0 and the enzyme showed amidolytic activity, since it hydrolysed the kallikrein synthetic substrate d-Pro-Phe-Arg-p-nitroanilide. The activity of sK1 upon rat plasma kininogen was strongly inhibited by the serine proteinase inhibitors phenylmethanesulfonyl fluoride, aprotinin or soybean trypsin inhibitor, but not by ethylenediaminetetraacetic acid or sodium tetrathionate. The molecular mass of sK1, as determined by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, was 66 kDa and the pI value, estimated by analytical chromatofocusing, was 4·2. Physical and chemical properties suggest that sK1 is a serine proteinase of the kallikrein family. Evidence is presented which suggests that sK1 is a component of the tegumental surface of the parasite and the levels of its activity in the male adult worm are approximately 21 times higher than those in the female adult worm. The intravenous injection of 3 μg of sK1 into an anaesthetized rat induced a drastic reduction in the arterial blood pressure of the animal. This effect lasted for about 1 min, and was followed by a progressive recovery of the arterial pressure. Neither bradycardia nor cardiac arrhythmias were noticed, suggesting a peripheral vasodilation effect. The presence of sK1 on the surface of adult male worms could play an important role in the wandering capacity of coupled worms into the visceral vasculature of the host.

A modification of the ‘floating raft’ method of in vitro penetration study is described. This allows replication of results and additional conveniences in operation. Filter paper membranes have shown characteristics similar to those of skin and have been used for quantitative studies. A method of digesting penetrating larvae from skin has been used to reduce the variability inherent in whole skin.Neither chloroform soluble skin products nor gravity were found to have a significant effect on the number of larvae entering membranes.Exsheathment of larvae was not essential prior to penetration and no specific stimuli for exsheathment appear to be necessary. Scanning electron micrographs have shown that entry may be by either hair follicles or desquamations of the stratum corneum.No evidence of enzymic activity during the invasion process has been found so far and the results suggest that a mechanical rather than chemical system obtains.

Pasteuria penetrans is a Gram-positive endospore-producing bacterium that is a parasite of root-knot nematodes. Attachment of endospores to the cuticle of the nematode is the first stage in the infection process. Western blot analysis with monoclonal and polyclonal antibodies that recognize the 30 kDa heparin-binding domain (HBD) and the 45 kDa gelatin-binding domain (GBD) fragments of human fibronectin (Fn) revealed a series of polypeptides of approximately 40, 45 and 55 kDa present in crude cuticle extracts of Meloidogyne javanica 2nd-stage juveniles. The results suggest that the structure of the nematode fibronectin is different to the fibronectins so far characterized. Pre-treatment of endospores of Pasteuria with either the HBD or the GBD was found to inhibit binding to the nematode cuticle. The larger GBD fragment was the most effective at blocking adhesion. Pre-treatment of the GBD fragment with gelatin prevented the GBD fragment from inhibiting endospore attachment to the nematode cuticle.

The mode of action of suramin upon Brugia pahangi has been investigated in vivo and in vitro. The drug was without effect on the glycolytic activity of worms in vitro at 2 × 10−4 M. The lack of effect was correlated with the failure of [14C]suramin to penetrate the worms in vitro. Suramin bound to the surface of worms in vitro presumably by virtue of its polyanionic nature. B. pahangi adults ingested [14C]suramin in vivo but no reduction in the rate of lactate production, of glucose utilization or in the rates of uptake of [14C]glucose, [14C]leucine or [14C]adenosine was observed in worms recovered from jirds between weeks 1 and 5 following 4 daily doses of suramin at 50 mg/kg given intraperitoneally. Worm death occurred between weeks 5 and 7 but this delayed drug effect was not the result of a progressive accumulation of suramin in the worms. Ultrastructural changes were observed in the intestinal epithelium of worms from suramin-treated jirds and parallel observations on worms exposed to Trypan blue in vivo suggest that both polyanionic compounds are restricted to the intestinal lumen of the worms. The evidence presented is consistent with the concept that, in B. pahangi, suramin acts at the surface of the intestinal epithelium and not by primarily inhibiting glucose catabolism or inhibiting phagosome and lysosome fusion as previously demonstrated for bloodstream trypanosomes and mammalian macrophages, respectively

The Trypanosoma brucei metacyclic population in the salivary glands of the tsetse fly displays a characteristic set of variable antigen types (VATs) which represents only a restricted part of the parasite's total VAT repertoire. After introduction into the mammalian host by fly bite, the metacyclics transform into bloodstream forms which retain expression of the metacyclic VATs. Specific antibodies, both polyvalent and monoclonal, have been used to neutralize separately 4 individual VATs from metacyclic populations. Control experiments and visual observation confirmed lysis of each VAT. On injection of the surviving trypanosomes, after washing, into mice each neutralized VAT was nevertheless expressed within a few days. Simultaneous neutralization of 2 metacyclic VATs which usually switch to one another in bloodstream infections did not prevent expression of either on subsequent injection into mice. Expression of neutralized VATs was not influenced by the antigenic composition of the population originally ingested by the tsetse fly. Metacyclic forms and their immediate successors thus appear to switch rapidly to expression of other metacyclic VATs in bloodstream populations.

Density-dependence in worm establishment, numbers, biomass and larval production were examined in primary infections of 0, 10, 40, 80 and 160 larvae of the lung nematode, Rhabdias bufonis in the common toad, Bufo bufo. The infection procedure established 4 non-overlapping levels of infection which persisted until 6 weeks post-infection (p.i.), after which there was an overall decline up to 12 weeks p.i. Worm numbers had no direct effect on adult worm survival but temporal changes in worm weight were density-dependent. Adult worm establishment in the lungs declined significantly as the numbers of worms in the lungs increased. At the lowest exposure dose, 86% of the larvae administered reached maturity in the lungs while at the highest, only 37% did so. Also, the numbers of immature larvae outside the lungs increased as adult worm numbers increased. Both features provide evidence for a threshold limit to the numbers of worms maturing in the lungs. Worm numbers also affected larval output per host and per capita fecundity. A significant positive relationship between per capita fecundity and per capita worm weight suggested that density-dependence acted primarily to constrain the growth of individual worms. Finally, the constraints imposed on worm growth and fecundity were apparently relaxed when worm density decreased, providing evidence for density-dependent flexibility in per capita fecundity. Density-dependence in worm establishment and per capita fecundity are mechanisms which may potentially regulate this host–parasite interaction in the field. Both mechanisms may be functionally related to physical space limitations in the lungs, within which worms must compete for finite nutrients.

Heliciculture is an excellent alternative to obtain edible snails but its viability is seriously threatened by pathogens. A parasitological survey was conducted in 3 mixed system-based heliciculture farms in Galicia (NW Spain), with the species Tetrahymena rostrata, Tetrahymena limacis, Tetratrichomonas limacis, Cryptobia helicogenae, Brachylaima aspersae (metacercariae and sporocysts), Alloionema appendiculatum, Nemhelix bakeri, and Riccardoella limacum being commonly found infecting Helix aspersa aspersa (petit-gris) snails. With the exception of C. helicogenae, N. bakeri, and B. aspersae sporocysts, all species were also detected in Helix aspersa maxima (gros-gris) snails, although generally with lower parameters. Most monoxenous infections, and consequently multiple parasitism, exhibited a rising trend during the first 2 months of intensive mating, with tendencies being slowed down or even reversed during the third month as a result of accumulated mortality and a sampling-derived reduction in host density. No parasites were vertically transmitted and infections were initially acquired from invading gastropod and micromammal reservoirs during fattening. Finally, artificial hibernation reduced significantly the prevalence of most species. These results confirm the importance of parasites in heliciculture and emphasize the need to prevent the entry of wild reservoirs into the farms and to rapidly remove the carcasses of dead snails from the reproduction units and fattening pens.

Clinical data on the use of artesunate combined with mefloquine in a variety of treatment regimens and parasite loads in Thailand were modelled on the basis of experimentally determined pharmacokinetic data. The model assumed no pharmacodynamic interaction between artesunate and mefloquine, but that the parasites were already resistant to mefloquine. Predictions of the model accorded well with the data. In particular, in accordance with clinical observations, the model showed that monotherapy with either drug failed to cure at moderate parasitaemia, yet such patients could be treated effectively with the combination of 3 days of artesunate + mefloquine. For high levels of parasitaemia, 5 days of artesunate+mefloquine were needed. Simulations were also performed for situations of lower resistance to mefloquine and for the immune human populations found in Africa. The importance of mathematical modelling of combination therapy is borne out by this study and suggests its wider application for other drug combinations.

We suggest a new mechanism for the maintenance of specificity of the association between the entomopathogenic nematode Steinernema scapterisci and its symbiotic bacteria. We evaluated the development and reproduction of infective and non-infective juvenile S. scapterisci in monoxenic combinations with its symbiotic bacteria, Xenorhabdus sp. ‘S’ and with the bacterial symbiont of Steinernema carpocapsae and Steinernema riobravis. Although development of non-infective stages occurred on all Xenorhabdus spp., the development of infective juveniles to the 4th stage (‘dauer’ recovery) was significantly delayed and reduced with X. nematophilus and Xenorhabdus sp. ‘R’, the bacterial symbionts of S. carpocapsae and S. riobravis, respectively. ‘Dauer’ recovery improved significantly when the cultures of X. nematophilus and Xenorhabdus sp. ‘R’ were supplemented with cell-free filtrates from Xenorhabdus sp. ‘S’. The infective juvenile S. scapterisci produced in all 3 cultures were virulent to Galleria mellonella larvae, confirming successful retention of Xenorhabdus from other steinernematids in their intestine. In fact, S. scapterisci infective juveniles containing X. nematophilus or Xenorhabdus sp. ‘R’ were more virulent to G. mellonella than those containing their natural symbiont, Xenorhabdus sp. ‘S’. We believe that this is the first demonstration of the symbiont-specific exit of infective juveniles from the ‘dauer’ phase which represents the finest level of specificity of bacteria–nematode association. This is also the first report of successful isolation of the natural symbiont of S. scapterisci.

The population of malaria-causing parasites is characterized by great genetic diversity. Knowledge of the polymorphism generation mechanism is a central issue for developing effective vaccines against malaria and understanding the parasite population structure. Plasmodium vivax genetic diversity has been explained in terms of two major factors: natural selection and intragenic recombination. A modular organization was found within P. vivax Duffy binding protein in the present work. Four Colombian isolates have identical sequences to Salvador-1 strain amongst dpb regions III–VI analysed, suggesting a high identity between Central and South American isolates. Geographically clustered sectors, corresponding to cysteine-rich regions (II and VI), show a high sequence diversity that could reflect a possible immune response evasion mechanism; both positive and negative selection were detected in these regions. In contrast, other dbp gene regions display a non-geographical clustering pattern, lower sequence diversity and predominant negative selective pressure. Recombination was homogeneously detected all along the molecule. These findings suggest that diversification vs. homogenizing forces, drive dbp gene evolution and determine its mosaic region organization.